ClC-3 promotes angiotensin II-induced reactive oxygen species production in endothelial cells by facilitating Nox2 NADPH oxidase complex formation
ClC-3 promotes angiotensin II-induced reactive oxygen species production in endothelial cells by facilitating Nox2 NADPH oxidase complex formation
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ClC-3 通过促进 Nox2 NADPH 氧化酶复合物的形成，促进血管紧张素 II 诱导的内皮细胞活性氧的产生
DOI：
10.1038/s41401-018-0072-0
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发表时间：
2018-11-01
影响因子：
8.2
通讯作者：
Du, Yan-hua
中科院分区：
文献类型：
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作者：
Liang, Guo-zheng;Cheng, Li-min;Du, Yan-hua
最近的证据表明，ClC-3，Cl-通道或Cl-/H+反向转运蛋白的ClC家族的成员，在NADPH氧化酶衍生的活性氧（ROS）的产生中起着关键作用。然而，其背后的机制仍不清楚。在这项研究中，我们研究了ClC-3对内皮细胞中NADPH氧化酶激活和ROS产生的影响和机制。血管紧张素II（Ang II，1 μ mol/L）处理显著提高培养的人脐静脉内皮细胞（HUVECs）中ClC-3的表达。此外，血管紧张素II治疗增加ROS的产生和NADPH氧化酶的活性，可以显着抑制敲低的ClC-3的效果，并进一步增强过表达的ClC-3。SA-β-半乳糖苷酶染色显示，ClC-3沉默消除Ang II诱导的HUVEC衰老，而ClC-3过表达引起相反的效果。我们进一步表明，血管紧张素II治疗增加了p47 phox和p67 phox从胞质到膜的易位，伴随着Nox 2和p22 phox表达的升高，这是显着减弱敲低ClC-3和增强过表达ClC-3。此外，ClC-3的过表达增加了血管紧张素II诱导的p47 phox和p38 MAPK的磷酸化。用p38抑制剂SB 203580预处理可消除ClC-3过表达诱导的p47phox磷酸化增加，以及NADPH氧化酶活性和ROS产生。我们的研究结果表明，ClC-3作为一个积极的调节剂，血管紧张素II诱导的NADPH氧化酶激活和ROS的生产，可能通过促进Nox 2/p22 phox的表达和p38 MAPK依赖的p47 phox/p67 phox膜转位，然后增加Nox 2 NADPH氧化酶复合物的形成。
Recent evidence suggests that ClC-3, a member of the ClC family of Cl- channels or Cl-/H+ antiporters, plays a critical role in NADPH oxidase-derived reactive oxygen species (ROS) generation. However, the underling mechanisms remain unclear. In this study we investigated the effects and mechanisms of ClC-3 on NADPH oxidase activation and ROS generation in endothelial cells. Treatment with angiotensin II (Ang II, 1 mu mol/L) significantly elevated ClC-3 expression in cultured human umbilical vein endothelial cells (HUVECs). Furthermore, Ang II treatment increased ROS production and NADPH oxidase activity, an effect that could be significantly inhibited by knockdown of ClC-3, and further enhanced by overexpression of ClC-3. SA-beta-galactosidase staining showed that ClC-3 silencing abolished Ang II-induced HUVEC senescence, whereas ClC-3 overexpression caused the opposite effects. We further showed that Ang II treatment increased the translocation of p47phox and p67phox from the cytosol to membrane, accompanied by elevated Nox2 and p22phox expression, which was significantly attenuated by knockdown of ClC-3 and potentiated by overexpression of ClC-3. Moreover, overexpression of ClC-3 increased Ang II-induced phosphorylation of p47phox and p38 MAPK in HUVECs. Pretreatment with a p38 inhibitor SB203580 abolished ClC-3 overexpression-induced increase in p47phox phosphorylation, as well as NADPH oxidase activity and ROS generation. Our results demonstrate that ClC-3 acts as a positive regulator of Ang II-induced NADPH oxidase activation and ROS production in endothelial cells, possibly via promoting both Nox2/p22phox expression and p38 MAPK-dependent p47phox/p67phox membrane translocation, then increasing Nox2 NADPH oxidase complex formation.